Nanoencapsulation as a General Solution for Lyophilization of Labile Substrates

Area of Science:

• Biotechnology
• Materials Science
• Protein Chemistry

Background:

• Proteins are nanoscale macromolecules crucial for biological functions.
• Current nanoparticle excipients often fail to protect proteins during lyophilization due to denaturation or lack of structural integrity.
• A novel approach is needed to preserve protein structure and function during freeze-drying.

Purpose of the Study:

• To investigate the potential of a nanoscale, thermostable exoshell (tES) as a protective excipient for protein macromolecules during lyophilization.
• To evaluate the ability of tES to prevent protein aggregation and denaturation throughout the freeze-drying process.
• To assess the recovery and retained activity of encapsulated proteins after reconstitution.

Main Methods:

• Systematic investigation of tES properties, including secondary structure and homogeneity, during lyophilization.
• Encapsulation of horseradish peroxidase (HRP) as a model protein substrate within tES nanoparticles using charge complementation and pH-mediated assembly.
• Time-course experiments to compare the activity of unprotected HRP versus tES-encapsulated HRP after lyophilized storage at various temperatures.

Main Results:

• tES demonstrated near 100% recovery after aqueous reconstitution, maintaining structural integrity.
• Encapsulation of HRP within 8 nm internal tES cavities was achieved via a simplified loading procedure.
• tES-encapsulated HRP retained 70% activity after lyophilized storage, a 14-fold improvement over unprotected HRP, which lost 95% activity.

Conclusions:

• Thermostable nanoencapsulation using tES is a novel and effective method for stabilizing functional macromolecules during lyophilization.
• This approach significantly enhances the long-term storage stability of labile proteins.
• tES nanoencapsulation represents a promising strategy for preserving protein activity in lyophilized formulations.